摘要
目的观察环孢素A(CsA)对人近端肾小管上皮细胞(HK-2)细胞转分化(EMT)及结缔组织生长因子(CTGF)表达的影响,探讨其致肾间质纤维化作用的可能机制。方法用相差显微镜观察不同浓度(0.5～25.0mg/L)CsA作用下HK-2细胞形态的变化;免疫荧光、RT-PCR、WesternBlot分别检测5mg/LCsA刺激细胞不同时间(24～72h)α-SMA在细胞内的表达和分布以及α-SMA、CTGFmRNA和蛋白的表达。结果显微镜下可见,HK-2细胞随CsA刺激浓度的增加由立方形铺路石样逐渐转变为梭形长条样。免疫荧光显示,随CsA刺激时间的延长细胞质中α-SMA表达逐渐增强。RT-PCR和WesternBlot显示,CsA呈时间依赖性上调α-SMA、CTGFmRNA和蛋白的表达,24h表达即明显增加,72h增加更为显著(P<0.01);CTGF与α-SMA表达的时间效应一致。结论环孢素A可上调CTGF的表达、诱导肾小管上皮细胞转分化,进而促进肾间质纤维化。
[Objective] To investigate the effects of cyclosporine A （CsA） on epithelial-to-mesenchymal transition （EMT） and connective tissue growth factor （CTGF） expression of human renal proximal tubular epithelial cells （HK-2） and to explore the possible mechanisms of renal interstitial fibrosis induced by CsA. [Methods] Different concentrations （0.5-25 mg/L） of CsA acted HK-2 cells for 48 hours and the same concentration （5 mg/L） of CsA acted at different time （48-72 hours）. Morphological changes were assessed by phase contrast microscopy. The mRNA and protein expression of α-SMA and CTGF were analysed by RT-PCR, indirect immunofluorescence and Western blot respectively. [Results] CsA induced striking morphological changes in HK-2 cells in a dose-dependent manner. Indirect immunofluoreseenee staining showed that expression of α-SMA could not be seen in control cells but upregulated and enhanced by 5 mg/L CsA in a time-dependent manner. CsA-induced EMT was also associated with increased expression of CTGF. The mRNA and protein expression of α-SMA and CTGF were both increased significandy in HK-2 cells induced by CsA in a time-dependent manner. [Conclusions] CsA up-regulates CTGF expression and induces EMT in renal tubular epithelial ceils, then leads to tubulo-interstitial fibrosis.
出处
《中国现代医学杂志》
CAS
CSCD
北大核心
2009年第1期72-75,80,共5页
China Journal of Modern Medicine
基金
福建省高效中西医结合基础重点实验室开放课题基金科研项目(No:zxy2008002)
2008年度陈可冀中西医结合发展基金项目(No:ckj2008006)
福建省卫生厅青年科研课题基金(No:2008-1-29)
福建省自然基金资助项目(No:2008J0278)
关键词
环孢素A
上皮细胞
转分化
结缔组织生长因子
cyclosporine A
epithelial cells
transition
connective tissue growth factor
